It is known that herpes simplex virus 2 (HSV-2) infects the genital mucosa and establishes a life-long infection in sensory ganglia. After a primary infection, HSV-2 may reactivate frequently, giving widespread genital lesions with considerable physical and psychological stress. A recent global estimate of HSV-2 infection concluded that 417 million people aged 15-49 years were infected in 2012, giving a world-wide prevalence of 11.3%. 19.2 million individuals were newly-infected 2012 (Looker, K. J., et al. 2015, PLoS One vol. 10, e114989). Furthermore, in newborns and in immunocompromised patients, HSV-2 infection can elicit severe and often fatal central nervous system infections.
As HSV-2 is the most common cause of genital ulcers globally, a major problem is that the infection strongly facilitates transmission of HIV. HSV-2 infection is associated with a three-fold increased risk of HIV acquisition among both men and women, implying that in areas of high HSV-2 prevalence a high proportion of HIV is attributable to HSV-2. This observation can be explained by the fact that local HSV-2 infection increases the number of HIV target cells, i.e. cervical CD4+ T-cells, in the genital mucosa. Furthermore, HSV-2 suppressive therapy with acyclovir reduced plasma HIV-1 RNA by 70% in dually infected women. The inventor concludes that an effective method for control of genital HSV-2 infection is of major public health importance. An efficacious vaccine will not only reduce genital lesions and other clinical manifestations of HSV-2 infection, but also reduce the risk of acquisition and transmission of HIV.
Human vaccine trials have been performed using HSV-2 glycoprotein B (gB-2) and/or glycoprotein D (gD-2) as antigens. The rationale has been that these proteins are essential for replication of HSV-2 and induce strong neutralizing antibody and T-cell responses in mice and humans. Despite these promising background data, results from randomized double-blind placebo-controlled multicenter trials have been unexpectedly discouraging. A gB-2- and gD-2-based vaccine was tested on 2,400 subjects and yielded only transient protection against infection (Corey et al., JAMA, 1999, vol. 282, 331-40). A prophylactic clinical study, including HSV-1 seronegative women (in total 8,323 subjects), was conducted between 2002 and 2010 in the U.S. and Canada. However, the results from this study revealed that the vaccine failed to prevent HSV-2 infection or disease (Belshe et al., 2012, N Engl J Med vol. 366, 34). These results clearly indicate a failure of previous vaccine strategies.
HSV-1 and HSV-2 are closely related viruses with an overall proteome amino acid sequence identity of >80%. The HSV-1 and HSV-2 envelope glycoproteins are strong inducers of the human antibody response. The high degree of similarity between these viruses implies that the envelope proteins, with the exception of glycoprotein G, contain several identical immunogenic regions and, therefore, elicit cross-reactive B- and T-cell responses. In other words, the immune responses elicited from an HSV-1 infection also recognise HSV-2-related antigens and vice versa. However, it seems clear that cross-reactive immune responses after an HSV-1 infection do not suffice to induce protective immunity against HSV-2 infection. This statement is based on the observation that a previous HSV-1 infection does not confer protection against acquisition of HSV-2. This notion makes sense from an evolutionary point of view, in that HSV-2 is mostly sexually transmitted, and usually infects persons at an older age than does HSV-1. Thus, HSV-2 has evolved to escape cross-reactive B- and T-cell responses induced by most of the HSV-1 glycoproteins. Therefore, the lack of protective properties of immune responses to cross-reactive proteins may explain both the failure of using the cross-reactive gB and gD proteins as vaccines, and why a prior HSV-1 infection is not protective for HSV-2 transmission.
The present invention is directed towards antigens of HSV-2 glycoprotein G (gG-2), and their use in vaccines and therapy, such as vaccines and therapy to prevent HSV-2 infection, and as antigens for use in the detection of anti-HSV-2 antibodies.
The gG-2 protein is a protein encoded by the US4 gene, which, in HSV-2 strain 333, comprises 2097 nucleotides including the stop codon. The US4 gene of HSV-2 strain 333 has the GenBank accession number EU018098.1, and its encoded amino acid sequence (i.e. the HSV-2 strain 333 gG-2 amino acid sequence) is presented herein as SEQ ID NO: 1. The nucleotide sequence of the complete HSV-2 strain 333 US4 gene is presented in SEQ ID NO: 13. The gG-2 protein is unique among the HSV proteins in that gG-2 is cleaved into one amino terminal secreted moiety (sgG-2), and one carboxy terminal intermediate, which includes a transmembrane region. The carboxy terminal intermediate is further processed to form mature, membrane-bound gG-2 (mgG-2), as described in detail below. SEQ ID NO: 2 shows the amino acid sequence of the mgG-2 protein of strain 333, consisting of 354 amino acids corresponding to amino acids 345-698 of SEQ ID NO:1.
It is known from the prior art (Liljeqvist et al., 1998, J Gen Virol., vol. 79, 1215; Marsden et al., 1998, J Med Virol., vol. 56, 79; Grabowska et al., 1999, J Gen Virol., vol. 80, 1789; Levi et al., 1996, Clin and Diagn. Lab Immunol., vol. 3, 265; Tunbäck, P. et al., 2000, J Gen Virol., vol. 81, 1033; Tunbäck, P. et al., 2005, J Gen Virol., vol. 86, 247) that the immunodominant antibody-binding regions of gG-1 (HSV-1 glycoprotein G) and mgG-2 are localized in the homologous regions of the proteins and not in the unique regions, see FIG. 3A-B. Despite the fact that some of these regions have high similarity between the two proteins, no cross-reactivity has been shown between human anti-gG-1 and anti-mgG-2 antibodies.
U.S. Pat. No. 5,665,537 claims that the unique region of gG-2 has desirable type-specific properties, and that antibodies which bind this region can be used for HSV serotyping. The document notes that antibody-binding regions are also localised in the homologous regions of gG-1 and mgG-2, and states that antibodies which bind these regions are cross-reactive with the two proteins.
In WO 98/003543 it is claimed that the antibody-binding region of human anti-mgG-2 antibodies, encompassing the 38 amino acids illustrated in FIG. 3D, is type-specific, and that this protein can be used for prophylactic, therapeutic and diagnostic uses relating to HSV-2 infection. However, this prior art document states that only the antibody-binding region of mgG-2 is important.
In WO 96/17938 glycoprotein D and G of HSV-2 and glycoprotein E of HSV-1 are used for vaccine purposes. It is disclosed that mgG-2 should be produced without both the transmembrane and the intracellular region.
WO 2010/135747 relates to proteins for use in a therapeutic and/or prophylactic HSV-2 vaccine. The application focuses on gD-2 and ICP-4, and presents limited data related to gG-2. Although GenBank accession number NP_044534.1 refers to entire gG-2 (Table 2, page 12) the protein specified in SEQ ID 38 (page 82) includes only sgG-2. Nothing is stated about the fact that gG-2 is cleaved into two proteins with different biochemical properties, i.e. sgG-2 and mgG-2. This document is completely silent in relation to mgG-2.
Görander, S. et al. disclose in Viruses, 2014, vol. 6, pages 4358-4372 (Anti-Glycoprotein G Antibodies of Herpes Simplex Virus 2 Contribute to Complete Protection after Vaccination in Mice and Induce Antibody-Dependent Cellular Cytotoxicity and Complement-Mediated Cytolysis), that anti-mgG-2 antibodies, elicited after vaccination of mice with full length mgG-2 and adjuvant, are important for the outcome after genital challenge with HSV-2.
Levi et al. disclose in Clinical and Diagnostic Laboratory Immunology, 1996, vol. 3, pages 265-269 (Peptide Sequences of Glycoprotein G-2 Discriminate between Herpes Simplex Virus Type 2 (HSV-2) and HSV-1 Antibodies), that synthetic glycoprotein G peptides may be useful for HSV-2 serology based on peptides, or combinations of peptides and antigens.
WO 97/05488, of Chiron Corporation, discloses a method of detecting and diagnosing herpes simplex virus (HSV) infection.
In view of shortcomings with vaccines against HSV-2 infections tested so far, there is a medical need for a new vaccine strategy, with efficacy against HSV-2 infections, or diseases related thereto.